A viral satellite DNA vector-induced transcriptional gene silencing via DNA methylation of gene promoter in Nicotiana benthamiana

Zheng Ju; Lei Wang; Dongyan Cao; Jinhua Zuo; Hongliang Zhu; Daqi Fu; Yunbo Luo; Benzhong Zhu

doi:10.1016/j.virusres.2016.07.005

A viral satellite DNA vector-induced transcriptional gene silencing via DNA methylation of gene promoter in Nicotiana benthamiana

Virus Res. 2016 Sep 2:223:99-107. doi: 10.1016/j.virusres.2016.07.005. Epub 2016 Jul 12.

Authors

Zheng Ju¹, Lei Wang², Dongyan Cao³, Jinhua Zuo⁴, Hongliang Zhu⁵, Daqi Fu⁶, Yunbo Luo⁷, Benzhong Zhu⁸

Affiliations

¹ College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: zhengju2016@126.com.
² College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: lwang2015@126.com.
³ College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: caody2012@126.com.
⁴ National Engineering Research Center for Vegetables, Beijing Academy of Agriculture and Forestry Sciences, Beijing 100097, China. Electronic address: zuojinhua@126.com.
⁵ College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: hlzhu@cau.edu.cn.
⁶ College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: daqifu@126.com.
⁷ College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: lyb@cau.edu.cn.
⁸ College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: zbz@cau.edu.cn.

PMID: 27422476
DOI: 10.1016/j.virusres.2016.07.005

Abstract

Virus-induced gene silencing (VIGS) has been widely used for plant functional genomics study at the post-transcriptional level using various DNA or RNA viral vectors. However, while virus-induced transcriptional gene silencing (VITGS) via DNA methylation of gene promoter was achieved using several plant RNA viral vectors, it has not yet been done using a satellite DNA viral vector. In this study, a viral satellite DNA associated with tomato yellow leaf curl China virus (TYLCCNV), which has been modified as a VIGS vector in previous research, was developed as a VITGS vector. Firstly, the viral satellite DNA VIGS vector was further optimized to a more convenient p1.7A+2mβ vector with high silencing efficiency of the phytoene desaturase (PDS) gene in Nicotiana benthamiana plants. Secondly, the constructed VITGS vector (TYLCCNV:35S), which carried a portion of the cauliflower mosaic virus 35S promoter, could successfully induce heritable transcriptional gene silencing (TGS) of the green fluorescent protein (GFP) gene in the 35S-GFP transgenic N. benthamiana line 16c plants. Moreover, bisulfite sequencing results revealed higher methylated cytosine residues at CG, CHG and CHH sites of the 35S promoter sequence in TYLCCNV:35S-inoculated plants than in TYLCCNV-inoculated line 16c plants (control). Overall, these results demonstrated that the viral satellite DNA vector could be used as an effective VITGS vector to study DNA methylation in plant genomes.

Keywords: Bisulfite sequencing; DNA methylation; Green fluorescent protein; Satellite DNA virus; Tomato yellow leaf curl China virus; Virus-induced transcriptional gene silencing.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

DNA Methylation*
DNA Viruses / genetics
DNA, Satellite*
DNA, Viral*
Gene Expression
Gene Order
Gene Silencing*
Genes, Reporter
Genetic Vectors / genetics*
Host-Pathogen Interactions / genetics
Nicotiana / genetics*
Nicotiana / virology*
Phenotype
Plant Diseases / genetics
Plant Diseases / virology
Plants, Genetically Modified
Promoter Regions, Genetic*

Substances

DNA, Satellite
DNA, Viral